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United States Patent | 6,001,635 |
Inoue ,   et al. | December 14, 1999 |
A strain of Sphingobacterium multivorum, mOL12-4s, is disclosed which produces deaminoneuraminidase in high yields. The deaminoneuraminidase produced by mOL12-4s does not act on the ketosidic linkages of N-acetylneuraminic acid or N-glycolylneuraminic acid containing complex carbohydrates, but only on ketosidic linkages of deaminoneuraminic acid containing complex carbohydrates.
Inventors: | Inoue; Yasuo (Taipei, TW); Inoue; Sadako (Taipei, TW); Kitajima; Ken (Nagoya, JP) |
Assignee: | Seikagaku Corporation (Tokyo, JP) |
Appl. No.: | 130929 |
Filed: | August 7, 1998 |
Jun 28, 1994[JP] | 6-146820 |
Current U.S. Class: | 435/252.1 |
Intern'l Class: | C12N 001/20 |
Field of Search: | 435/252.1 |
5449615 | Sep., 1995 | Li et al. | 435/200. |
5679557 | Oct., 1997 | Ito et al. | 435/156. |
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TABLE 1 ______________________________________ KDNase activity KDNase per cells Added Number of activity (milliunit/10.sup.10 substance cells (milliunit) cells) ______________________________________ Before 6.1 .times. 10.sup.10 [1] 20 [1] 3.3 [1] addition Glc 1.7 .times. 10.sup.12 [28] 80 [4] 0.47 [0.14] KDN-OS 1.5 .times. 10.sup.12 [25] 7500 [380] 50 [15] crude 5.7 .times. 10.sup.12 [93] 23000 [1200] 40 [12] KDN-OS KDN 3.6 .times. 10.sup.10 [0.59] 38 [1.9] 11 [3.3] Neu5Ac 2.5 .times. 10.sup.10 [0.41] 15 [0.75] 6.0 [1.8] Oligo- 3.0 .times. 10.sup.10 [0.49] 12 [0.60] 4.0 [1.2] Neu5Ac ______________________________________
TABLE 2 ______________________________________ Purifi- Specific cation Activity Protein activity Yield degree Fraction (unit) (mg) (unit/mg) (%) (fold) ______________________________________ cell-disrupted 153 207 0.737 100 1.0 solution 50-70% 113 103 1.11 74.2 1.5 (NH.sub.4).sub.2 SO.sub.4 fraction 1st Sephacryl 76.1 57.3 1.33 49.8 1.8 S-200 2nd Sephacryl 57.4 26.6 2.16 37.6 2.9 S-200 KDN-gp agarose 52.2 0.410 127 34.2 173 adsorbed fraction ______________________________________
TABLE 3 __________________________________________________________________________ Presence or absence of Complex carbohydrate or carbohydrate Linkage cleavage __________________________________________________________________________ 4-MU-KDN + KDN dimer KDN.alpha.2.fwdarw.8KD + KDN-containing double strand N-type sugar chain KDN.alpha.2.fwdarw.3Gal + KDN oligosaccharide alcohol KDN.alpha.2.fwdarw.8KDN, KDN.alpha.2.fwdarw. 3Gal + KDN.alpha.2.fwdarw.6GalNAcol KDN-containing glycoprotein KDN.alpha.2.fwdarw.8KDN, KDN.alpha.2.fwdarw. 3Gal + KDN.alpha.2.fwdarw.6GalNAc KDN-containing ganglioside GM3 KDN.alpha.2.fwdarw.3Ga1 + 4-MU-Neu5Ac Neu5Ac dimer Neu5Ac.alpha.2.fwdarw.8Neu5Ac - Neu5Gc dimer Neu5Gc.alpha.2.fwdarw.8Neu5Gc - Neu5Ac lactose Neu5Ac.alpha.2.fwdarw.3(6)Gal - Neu5Ac-containing double strand N-type sugar chain Neu5Ac.alpha.2.fwdarw .3Gal - Neu5Ac.alpha.2.fwdarw.6(Gal.beta.1.fwdarw.3)GalNAcol Neu5Ac.alpha.2.fwda rw.6GalNAcol - human transferrin Neu5Ac.alpha.2.fwdarw.6Gal - fetal bovine serum fetuin Neu5Ac.alpha.2.fwdarw.3(6)Gal - swine submandibular gland mucin Neu5Gc.alpha.2.fwdarw.6GalNAc - toad ovum jelly glycoprotein Neu5Ac.alpha.2.fwdarw.6GalNAc - colominic acid (.fwdarw.8Neu5Ac.al pha.2.fwdarw.).sub.n - lake trout polysialoglycoprotein (.fwdarw.8Neu5Ac.alpha.2.fwdarw.).sub.n - rainbow trout polysialoglycoprotein (.fwdarw.8Neu5Gc.alpha.2.fwdarw.).su b.n - arctic char polysialoglycoprotein (.fwdarw.8Neu5Ac.alpha.2.fwdarw.,.fwda rw.8Neu5Gc.alpha.2.fwdarw.).sub.n - Neu5Ac-containing ganglioside GM3 Neu5Ac.alpha.2.fwdarw.3Gal - Neu5Ac-containing ganglioside GM1 Neu5Ac.alpha.2.fwdarw.3(GalNAc.beta.1. fwdarw.4)Gal - __________________________________________________________________________